Major histocompatibility complex – antigen processing and presentation

Question 1

What are the characteristics of immature B and T cells

Answer 1

Immature B and T cells are small and motile and are non-phagocytic

They are morphologically identical – they do however have a unique receptor which can be detected using monoclonal antibodies

Question 2

What stage of the cell cycle are naive lymphocytes ub

Answer 2

G0

(they are quiescent) - they can be induced into the cell cycle via Ag stimulation

Question 3

What happens as they progress through the cell cycle

Answer 3

They proliferate and differentiate

Effector cells (mediate immune function and are short-lived)

Memory cells (enhance response to previously seen Ag and are life-long)

Question 4

types of immunogens and their potency

Answer 4

Proteins are the most potent immunogen

Polysaccharides are also potent but not as potent

Lipids/nucleic acids must be linked to proteins or polysaccharides to be immunogenic

Question 5

What are the criteria for good immunogens

Answer 5

Foreignness – must be recognised as “non-self”

Molecular size (>100kDa best 5-10 kDa poor)

Chemical composition / complexity

Homopolymers (lack of immunogenicity regardless of size)

Copolymers >2 different amino acids / sugars are good

Ability to be recognised

Question 6

How do B cells recognise antigens

Answer 6

B cells recognise soluble (or processed) antigen via B cell receptor (membrane bound antibody) - epitopes must be accessible as Ag is free in solution

Naïve B cells encounter Ag – activated – proliferate and differentiate into Ab secreting plasma (effector) cells

Roughly 10% of B cells mature and exit the bone marrow

They will die in 2 days if no Ag is encountered

Professional antigen presenting cells – soluble Ag bound to B cell receptor is internalised, processed and displayed at B cell surface coupled to MHC-II molecules

Question 7

How do T cells recognise antigens

Answer 7

React with internally processed Ag – a peptide linked with an MHC molecule on the surface of antigen presenting cells (APCs) - can also be altered self

T cells must be shown a processed antigen to be activated

2 classes (broadly) of T cells

Cytotoxic T cells (CD8+) - killer cells involved with Ag-specific cell killing of altered self cells

T helper cells (CD4+) - secrete cytokines and coordinate the immune response

Question 8

What is an immunological synapse and what is its function

Answer 8

a specialized cell-to-cell junction where immune cells, like T cells or NK cells, interact with other cells, such as antigen-presenting cells (APCs)

Question 9

What interacts with each other at an immunological synapse

Answer 9

Engagement with the Ag associated MHC on the APC with the T cell receptor and CD4 or CD8 to distinguish MHC-II and MHC-I

MHC-II interacts with CD4 and MHC-I interacts with the CD8

Co-stimulation CD80 / CD86 on APC with CD28 are required for full T cell activation

CTLA-4 engagement supresses response (negative feedback – acts as an “off” signal)

Question 10

What are cluster of differentiation molecules (CD molecules)

Answer 10

Categorisation scheme of cell surface membrane (glycoproteins)

> 371 CD molecules

Question 11

What do CD molecules do

Answer 11

Allow for the identification of leukocyte subsets

Fluorescent conjugated CD specific antibodies label cells that express a particular CD molecule (immunophenotype)

CD molecules have a defined function

Question 12

What are important CD molecules

Answer 12

CD3 – expressed on T cells and is the signalling component part of the T cell receptor complex

CD4 – co-receptor for MHC-II by T helper cells

CD8 – co-receptor for MHC-I expressed by T cytotoxic cells

CD45 – common leukocyte antigen

T helper cells have the immunophenotype : CD45+CD3+CD4+ (this defines a T cell and the CD4+ part defines the class)

T cytotoxic cells – CD45+CD3+CD8+

CD19 – B cell receptor component

CD25 – interleukin-2 (IL-2) receptor (this is up-regulated during activation)

Question 13

MHC characteristics

Answer 13

Are polymorphic antigen presenting proteins

They are markers of “self”

MHC bind processed Ag at the cell surface for T cells to recognise

T cells require the presentation of processed antigen for activation

Question 14

What are the biological functions of MHC molecules

Answer 14

MHC-I – present on almost all nucleated cells and present endogenous peptide to T cytotoxic cells (CD8+)

MHC-II – present on APC and present exogenous peptide to T helper cells (CD4+)

Structurally are fairly homologous

Question 15

What are the characteristics of MHC-I

Answer 15

Present on all nucleated cells

Present endogenous peptide which includes self, altered self (malignant) and viral (allows infected cells to be detected)

MHC-I binds to CD8 molecule on cytotoxic cells

Basic structure

Two subunits

The alpha-chain has 3 extra cellular domains α1, α2 and α3

Also has a transmembrane segment and cytoplasmic tail

A β2-microglubulin – binding cleft α1-α2 can bind peptide 8-13 residues (usually 9 residues)

Question 16

What are the characteristics of MHC-II

Answer 16

Present on APC

Present exogenous peptide from bacteria or yeast etc

MHC-II binds to CD4 molecules on T helper cells – this produces cytokine, inflammatory response and stimulates Ig class switching in B cells

Basic structure

Two subunits (α and β chains)

Alpha and beta-chains have 2 extra-cellular domains α1 / α2 and β1 / β2 respectively

They also have transmembrane segments and cytoplasmic tails

Binding cleft α1-β2 can bind peptide 12-18 residues (open ended)

Question 17

MHC polymorphisms

Answer 17

Peptide binding cleft of MHC molecules is polymorphic

Alters the peptide / MHC affinity which creates stronger or weaker interactions (based on how the peptide sits in the cleft)

Peptide / MHC association is degenerate

Question 18

What are the interactions within the cleft of a MHC molecule

Answer 18

Dipole-dipole

Hydrogen bonding

London dispersion

Hydrophobic

Residue alterations may alter complementary sticky patches at key areas

Question 19

How do MHC and disease susceptibility link

Answer 19

Certain MHC alleles are associated with increased risk of certain diseases which include viral infections and autoimmune diseases

Examples :

HLA-DR4 – associated with rheumatoid arthritis and type-1 diabetes

HLA-DQ2 – associated with increased risk of coeliac disease

HLA-DQB1 – associated with narcolepsy

HLA-B57 – associated with greater HIV control (slower progression)

Question 20

What are examples of this in organisms

Answer 20

A lack of outbreeding in cheetah populations has meant that there is limited MHC polymorphisms, making cheetahs more susceptible to viral diseases

Tasmanian devils are able to spread facial tumour – rare example of transmissible tumour through biting

Question 21

What are the HLA genes

Answer 21

HLA genes, or Human Leukocyte Antigens, are a set of genes that play a critical role in the immune system by helping it distinguish between the body’s own cells and foreign invaders. They are part of the Major Histocompatibility Complex (MHC)

The MHC is coded by the human leukocyte antigen (HLA) complex

Question 22

What are the characteristics of the HLA locus

Answer 22

The HLA locus is one of the most genetically diverse parts of the human genome

Most variable regions are HLA-B and HLA-DRB1

Question 23

What does the HLA locus contain

Answer 23

MHC-I genes (A,B and C)

MHC-II (DP,DQ and DR)

MHC-III (some complement proteins and cytokines)

Question 24

What is the pattern of inheritance for HLA

Answer 24

HLA antigen are co-dominant meaning maternal and paternal alleles are expressed

MHC genetic diversity is inherited (no somatic recombination)

As MHC genes are so closely linked, the chance of genetic crossover is rare during meiosis

Class-I and class-II MHC genes are inherited together – haplotype

Question 25

Why is tissue typing important in transplantations

Answer 25

In organ or tissue transplantation MHC acts as an antigen Therefore the donor and recipient needs to be matched (as MHC is a marker of self) We each have 6 MHC alleles (HLA-A, HLA-B and HLA-C from each parent) We also have 6-8 MHC-II alleles (HLA-DP, one HLA-DQ and one or two HLA-DR from both parents) HLA-A, HLA-B or HLA-DR compatibility is assessed before donation Hyperacute rejection – pre-existing recipient antibodies (previous transfusion) Acute/chronic humoral rejection – recipient generates anti-donor MHC response after the transplantation

Question 26

How are antigens processed and presented

Answer 26

Antigen is presented at the cell surface by the MHC Intracellular processes cleave the protein into peptide fragments – these associate with the MHC and is presented in the plasma membrane 2 major peptide antigen process mechanisms can occur Endogenous antigens – processing occurs in all nucleated cells – couples peptide to MHC-I for CD8+ T cell presentation Exogenous antigen – processing occurs in APC – couples peptide with MHC-II for CD4+ T cell presentation

Question 27

How does exogenous antigen processing and presentation differ to endogenous processing and presentation

Answer 27

Arise from extracellular pathogens (bacteria etc) Phagocytosis of pathogen – degraded in phagosome by endopeptidase activity generates 12mer to 18mer peptide fragments In the ER the MHC-II molecule is complexed with the "invariant chain" This blocks binding at the binding cleft (MHC-II Ii) which prevents binding of endogenous peptide Invariant chain is cleaved creating MHC-II CLIP complex Fusing with phagosome – HLA-DM/DO mediates the release of CLIP – allows binding of peptide to MHC-II There are several crucial points of interaction These are called anchor points Position 4 = negative charge Position 9 = hydrophobic Degenerate peptide binding to MHC-II Negatively charged and hydrophobic anchor points are aligned Open ended MHC-II with peptide (looks like a hotdog in a bun)

Question 28

How does endogenous antigen processing and presentation differ to exogenous processing and presentation

Answer 28

Arise from intracellular pathogens (virus etc) Self or viral antigen – degraded in proteasome by endopeptidase which generates 8mer to 13mer peptide fragments MHC-I / peptide complex in ER – moved to cell surface by a vesicle Endogenous peptides enter the ER via the transport associated with antigen processing (TAP) protein MHC-I in ER is associated with calreticulin and tapasin proteins – both act as chaperones for peptide loading Once the peptide is loaded the complex is exported to the plasma membrane Every cell must have MHC-I as they all must be killable by CD8 Anchor points for MHC-I are better defined (set rules) Position 2 = charged (N-term) Position 9 = hydrophobic (C-term) Degenerate residues in between longer peptides (bulge) Degenerate binding to MHC-I Unlike MHC-II (open-ended) MHC-I is closed-ended Second (N-term) and last (C-term) residues of the peptide must be charged and hydrophobic (8mer and 9mer fit cleft perfectly) Large peptides (11mer and 13mer) bulge in the middle The larger they are the greater the steric hindrance with respect to TCR interaction

Question 29

How are non-peptide antigens processed and presented

Answer 29

Nucleic acids and lipids can also be immunogenic (to a lesser extent though) T cells react to lipids like mycolic acid through MHC like molecules CD1 is structurally similar to MHC-I but it functions in a similar way to MHC-II (exogenous antigen presentation)

Question 30

How are lipids presented by CD1

Answer 30

Hydrophobic residues within CD1 binding site (deeper narrower groves) interact with fatty-acid side chains (carbohydrates) polar regions is exposed to the extracellular fluid This is recognised by T cells